Gallbladder carcinoma (GBC) is one of the most common carcinomas of the biliary tract and is associated with aggressive malignancy and poor prognosis. Current therapeutic strategies, including surgery, radiotherapy, and chemotherapy, are not sufficient for the treatment of GBC, and new therapeutic strategies are urgently needed. The antitumor effects of oroxylin A (OrA), a natural flavonoid extracted from the dried roots of medicinal plants such as Scutellariae species (Radix Scutellariae), have been widely reported in various cancers. In this study, we first evaluated the antitumor activity and the underlying mechanism of action of OrA on GBC cells in vitro . Our results revealed that OrA significantly attenuated the proliferation, migration, and invasion of GBC cells, simultaneously promoting their apoptosis. Suppression of the phosphate on and tension homology deleted chromosome ten (PTEN)/phosphatidylinositol-3 kinase (PI3K)/protein kinase B (AKT) signaling pathway was found to be the underlying mechanism involved in the antitumor activity of OrA. In addition, experiments using a tumor xenograft mouse model confirmed the antitumor effects of OrA in vivo . Taken together, our findings indicate that OrA could be a potential antitumor agent for the prospective treatment of GBC.